The broad goals of this project are to understand the control of enzyme formation in the arginine biosynthetic pathway in Escherichia coli. Enzyme synthesis in the arginine pathway is regulated by a repressor protein (argR) which interacts at operator sites in the 6 arginine operons to repress transcription of arg mRNA. The purpose in this proposal is to purify the repressor protein and to study its mode of action and to identify and determine the nucleotide sequences of regulatory DNA in the arginine operons. Using cell-free synthesis of beta-galactosidase from a lambda DNA template carrying a fusion of lacZ to the argA promoter as an assay for arginine repressor activity, it was possible to partially purify the repressor protein. In order to increase the cellular concentration of repressor for the purpose of purifying this protein, studies are proposed to place the argR gene on a multicopy hybrid plasmid under the control of the lac promoter. Once pure, studies of the mechanism of repressor action in preventing transcription will be carried out. Isolation of the lambda phage carrying the argA-lacZ fusion has made it possible to identify two restruction fragments of 70 and 110 bases, respectively. These will be analyzed for nucleotide sequence determinations. Lambda phages carrying lac fusions to argE, argD and argG have also been constructed and these will be analyzed by restriction enzyme analysis to identify short segments of arg regulatory DNA suitable for sequence analysis.